Stable pharmaceutical compositions

ABSTRACT

Pharmaceutical compounds having the general formula: 
                         
or compounds having the formula:
 
                         
where R 1  and R 2  are independently H, branched or straight chain alkyl having from 1 to about 7 carbon atoms, substituted or unsubstituted aryl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aralkyl, substituted or unsubstituted alkylcycloalkyl, lower alkenyl or R 1  and R 2  together form part of a substituted or unsubstituted cycloalkyl having from about 4 of about 7 carbon atoms; where R 3  is a branched or straight chain alkyl having from 1 to about 7 carbon atoms, substituted or unsubstituted aryl, substituted or unsubstituted aralkyl, substituted or unsubstituted cycloalkyl, aralkyl, substituted or unsubstituted alkylcycloalkyl or a group having the formula (CH 2 ) n COOH where n is from 1 to about 7.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional application of U.S. patent applicationSer. No. 10/306,196 filed Nov. 27, 2002 now U.S. Pat. No. 6,821,974which is a continuation-in-part to patent application Ser. No.10/087,951 entitles Stable Pharmaceutical Compositions, filed Mar. 5,2002 now U.S. Pat. No. 6,825,196 in the name of Donald L. Barbeau.

BACKGROUND OF THE INVENTION

The present invention relates to both a novel method of stabilizinghydralazine hydrochloride in pharmaceutical preparations andpharmaceutical compositions containing stabilized hydralazine compoundshaving the general formula:

or compounds having the formula:

where R₁ and R₂ are independently H, branched straight chain alkylhaving from 1 to about 7 carbon atoms, substituted or unsubstitutedaryl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted aralkyl, substituted or unsubstituted alkylcycloalkyl,lower alkenyl or R₁ and R₂ together form part of a substituted orunsubstituted cycloalkyl having from about 4 to about 7 carbon atoms;where R₃ is a branched or straight chain alkyl having from 1 to about 7carbon atoms, substituted or unsubstituted aryl, substituted orunsubstituted aralkyl, substituted or unsubstituted cycloalkyl, aralkyl,substituted or unsubstituted alkylcycloalkyl or a group having theformula (CH₂)_(n)COOH where n is from 1 to about 7.

Alternatively, the present invention relates to pharmaceuticalcompositions containing compounds having the general formula:

or compounds having the formula:

where 1 where R₁ and R₂ are independently H, branched or straight chainalkyl having from 1 to about 7 carbon atoms, substituted orunsubstituted aryl, substituted or unsubstituted cycloalkyl, substitutedor unsubstituted aralkyl, substituted or unsubstituted alkylcycloalkyl,lower alkenyl or R₁ and R₂ together form part of a substituted orunsubstituted cycloalkyl having from about 4 of about 7 carbon atoms;where R₃ is a branched or straight chain alkyl having from 1 to about 7carbon atoms, substituted or unsubstituted aryl, substituted orunsubstituted aralkyl, substituted or unsubstituted cycloalkyl, aralkyl,substituted or unsubstituted alkylcycloalkyl or a group having theformula (CH₂)_(n)COOH where n is from 1 to about 7.

Hydralazine hydrochloride is a peripheral vasodilator discovered about50 years ago that exerts an antihypertensive effect directly on vascularsmooth muscle producing relaxation of muscle fibers resulting in adecrease in blood pressure. Hydralazine is extensively metabolized inthe body to products that are excreted predominantly in the urine, andundergoes N-acetylation, oxidation, hydroxylation, hydrazone formationand conjugation.

Commercially available in both oral and injectable dosage forms,hydralazine is used to lower blood pressure in hypertensive crisissituations and in patients requiring long-term management of theirhypertension after the crisis has abated. Hypertensive crisis is amedical emergency that requires immediate therapy for certain patientsin hospital emergency rooms, operating rooms and intensive care units.

Hydralazine is an artery specific direct peripheral vasodilator havingan onset of action between 10-30 minutes (10-20 minutes givenintravenously), a maximum hypotensive effect in 10-80 minutes andduration of action between 3-4 hours. Hydralazine is one of the fewinjectable antihypertensive drugs that maintain blood flow to kidneysduring hypertensive crisis, and the only one to increase blood flow toan already compromised kidney. Although the drug is approved foradministration of 20-40 mg doses, there are several clinical hazardsassociated with the currently available hydralazine formulation. First,the instability of the 20 mg/ml sterile solutions is a serious problemand has frequently caused removal of the product from the market by theFDA. Submicron particles appear in the hydralazine sterile injectionsolutions during storage for more than 6-12 months. Secondly, theconcentrated 20mg/ml dosage form of hydralazine is difficult toadminister accurately to patients at the small doses (3-5 mg) requiredto avoid “overshoot” hypotension. Consequently, these concentratedsolutions are generally diluted prior to use in the hospital.Unfortunately, dilution by hospital personnel in an attempt to reducethe administration problems risks alteration of the hydralazine product,metal contamination and generation of toxic substances. Commerciallyavailable hydralazine solutions discolor when inappropriately dilutedwith metal-containing or carbohydrate-containing diluents generallyfound in hospitals. The Food and Drug Administration (FDA)s labeling forthe currently available hydralazine formulation indicates thathydralazine should not be added to infusion solutions, and thathydralazine hydrochloride injections may discolor upon contact withmetal. The FDA further warns in the product labeling that discoloredsolutions should be discarded.

Hypertensive crisis is a life-threatening situation and includeshypertensive emergencies and hypertensive urgencies characterized byacute elevations in blood pressure, which must be brought under controlwithin hours. Over 60 million people in the United States suffer fromessential hypertension. About 1% of these people suffers fromhypertensive crisis and requires hospital-based acute care. Of thehypertensive crisis patients, 76% are “urgencies” and 24% “emergencies”with end-organ damage. Hypertensive urgencies are those situations inwhich it is desirable to reduce blood pressure quickly; however,hypertensive urgencies can be managed without requiring rapid,controlled reduction of blood pressure. Elevated blood pressure alone,in the absence of symptoms or progressive target organ damage rarelyrequires emergency treatment. Hypertensive urgencies are treated withoral antihypertensives such as nifedipine or clonidine, or withintravenous labetolol.

Hypertensive emergencies are characterized by acute elevations in bloodpressure (diastolic >110 to 120 mm Hg) which can potentially be lifethreatening and thus require rapid, controlled reduction of bloodpressure. Prompt pharmacologic therapy is indicated for those patientshaving Stage 2 (≧160/100 mm Hg) or Stage 3 (≧180/110 mm Hg) hypertensionwho have clinically manifested cardiovascular disease or target organdamage. The most commonly used antihypertensive agent is nitroprusside.Although hydralazine is already labeled for severe essentialhypertension when oral hydralazine cannot be given or when the need tolower blood pressure is urgent as in hypertensive crisis, it is notcurrently labeled for hypertensive emergencies when a patient presentswith emergent end-organ damage. As a patient's blood pressure is acutelyelevated, the patient experiences a dramatic decrease in blood flow tovital tissues such as the kidney and brain. The reduction in elevatedblood pressure in these patients through antihypertensive therapy isimportant because it minimizes ischemic damage resulting from reducedblood flow to these tissues. Examples of emergent end-organ damageinclude hypertensive encephalopathy, cerebral infarction, intracranialhemorrhage, myocardial ischemia, acute pulmonary edema, hypertensivenephropathy, hypertensive retinopathy and eclampsia. The goal of therapyin hypertensive emergencies is to reduce the mean arterial pressure byno more than 25 percent with two hours, then toward 160/110 mm Hg within2 to 6 hours avoiding excessive drops in pressure that may precipitateor aggravate renal, cerebral or coronary ischemia. Ultimately, the goalof therapy is to reduce the blood pressure to below 140/90 mm Hg.

Hydralazine hydrochloride is very unstable in all of the injectablepharmaceutical formulations currently commercially available. Continuinginstability problems with injectable hydralazine hydrochloride, forexample, have plagued pharmaceutical manufacturers for many years,forcing these companies to remove their injectable hydralazine productsfrom the marketplace. Although a shelf life of 12 months is currentlyrequired for FDA approval for injectable hydralazine hydrochloride, onlya few companies have been able to satisfy this requirement with adequatestability data. One such company, SoloPak Pharmaceuticals, Inc. was ableto meet the 12-month stability requirements for FDA approval; however,the company was not able to provide a drug product that was consistentlystable for more than 6 months.

In its injectable formulation, hydralazine forms small yellow-greenparticles following storage from 1 to about 2 months when hydralazine isstored at 40° C. and after from 6 to about 9 months storage at 25° C.Although the identification of the yellow-green particles has yet to beconfirmed, it is believed that the particles are insoluble polymericproducts formed during storage of hydralazine. It is believed thathydralazine hydrochloride undergoes degradation in stored sterileinjectable solutions to insoluble polymeric products due to the highlyreactive hydrazino group. Hydralazine hydrochloride also undergoesseveral pharmaceutically undesirable reactions such as chelation withmetal ions, oxidation, and pH-dependent decomposition. It is believedthat these reactions, which often cause discoloration of hydralazinecompositions, are also due to the highly reactive hydrazino group.

Kanazawa et al. [Chemical and Pharmaceutical Bulletin 34(4):1840-1842(1996)] report that during the storage of a prescription admixture ofpulverized hydralazine hydrochloride with cimetidine, a histamineH₂-receptor antagonist for duodenal ulcer, the initially uncoloredadmixture gradually turns to pale yellow. Kanazawa et al. further reportthat hydralazine hydrochloride undergoes degradation and discolorationwith cimetidine in aqueous solution to give1,1-di(phthalazin-3-yl)amine, 1,1-di(phthalazin-3-yl)hydrazine,1-amino-1,2, 2-tri-(phthalazin-3-yl)hydrazine, and1,1,2-tri(phthalazin-3-yl)hydrazine.

Alexander et al. [American Journal of Hospital Pharmacy 50:683-686(1993)] report that the degradation of hydralazine hydrochloridein a sugar-containing oral syrup was quite fast and was apparently afirst order process. The authors report that sugar (e.g., dextrose andfructose) reduces the stability of hydralazine hydrochlorideconsiderably. Their syrup containing maltitol normally increases thestability of drugs sensitive to the presence of sugars; however, thehydralazine formulation remained unstable at room temperature.

Lessen et al. [Journal of Pharmaceutical sciences 85(3): 326-329(1996)]report that the strength of hydralazine hydrochloride in 10 mg tabletscontaining starch as an excipient decreases significantly with time andproduced fluorescence at 414 nm. Lessen at al. report that, in additionto the usual hydralazine degradants such as phthalazone and phthalazine,these tablet compositions produced triazolophthalazine derivatives.

Hydralazine is known to chelate metal ions. Sinha and Motten[Biochemical and Biophysical Research Communications 105(3):1044-1051(1982)] report that hydralazine oxidizes rapidly in thepresence of oxygen and metal compounds such as Cu⁺², Fe⁺², and Fe⁺³through free radical intermediates much like other hydrazinederivatives.

Because of its reactivity toward metals, standard manufacturingrequirement for the preparation of bulk hydralazine solution or sterilefill solution is that neither should come into contact with any metalsurface including tanks, transfer lines or filling lines. Unfortunately,these precautions can be consistently enforced by the manufacturer onlyduring preparation and storage of the hydralazine solutions. Afterstorage of the hydralazine solutions, the handling of the hydralazinesolutions is no longer under their control.

Despite its unique pharmacologic properties as a hypertensive drug, thetherapeutic use of hydralazine hydrochloride has been limited by itsinstability during storage and difficulties in handling by medicalpersonnel. A stable hydralazine pharmaceutical composition that is moreeasily manufactured and does not degrade or produce particulate matterduring extended storage does not currently exist. Moreover, aninjectable hydralazine pharmaceutical formulation that is not adverselyaffected by conventional dilution techniques in the hospital does notcurrently exist. This, despite the fact that hydralazine was discoveredas an antihypertensive agent over 50 years ago. A stable hydralazinecomposition that could be manufactured more easily and stored forperiods of time greater than the current 12 month limit represents asignificant advance.

BRIEF SUMMARY OF THE INVENTION

This invention relates to a method of improving the stability of ahydralazine composition during manufacturing or storage comprisingcoupling an N-protecting group with hydralazine to produce the compoundhaving the formula:

or a compound having the formula:

where R₁ and R₂ are independently H, branched or straight chain alkylhaving from 1 to about 7 carbon atoms, substituted or unsubstitutedaryl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted aralkyl, substituted or unsubstituted alkylcycloalkyl,lower alkenyl or R₁ and R₂ together form part of a substituted orunsubstituted cycloalkyl having from about 4 to about 7 carbon atoms;where R₃ is a branched or straight chain alkyl having from 1 to about 7carbon atoms, substituted or unsubstituted aryl, substituted orunsubstituted aralkyl, substituted or unsubstituted cycloalkyl, aralkyl,substituted or unsubstituted alkylcycloalkyl or a group having theformula (CH₂)_(n)COOH where n is from 1 to about 7; and where saidN-protecting group is removed from said compound after manufacturing orstorage.

Alternatively, the present invention relates to pharmaceuticalcompositions containing hydralazine compounds having the generalformula:

where X is a group having the formula

or compounds having the general formula:

where X is a group having the formula

where R₁, R₂ and R₃ having the same meaning as described above.

In particular, the present invention relates to pharmaceuticalcompositions containing compounds having the formula:

or compounds having the formula:

where R₁ and R₂ are independently H, branched or straight chain alkylhaving from 1 to about 7 carbon atoms, substituted or unsubstitutedaryl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted aralkyl, substituted or unsubstituted alkylcycloalkyl,lower alkenyl or R₁ and R₂ together form part of a substituted orunsubstituted cycloalkyl having from about 4 of about 7 carbon atoms;where R₃ is a branched or straight chain alkyl having from 1 to about 7carbon atoms, substituted or unsubstituted aryl, substituted orunsubstituted aralkyl, substituted or unsubstituted cycloalkyl, aralkyl,substituted or unsubstituted alkylcycloalkyl or a group having theformula (CH₂)_(n)COOH where n is from 1 to about 7.

It is an object of the present invention to stabilize pharmaceuticalcompositions during manufacturing so that the hydralazine does not reactwith metal components of the manufacturing system.

It is a further object of the present invention to extend the shelf lifeof oral and injectable pharmaceutical compositions containinghydralazine significantly beyond 12 months storage and preferably beyond24 months storage.

It is a further object of the present invention to stabilize injectablepharmaceutical compositions containing hydralazine during storage andreduce the formation of submicron particles.

It is a further object of the present invention to stabilize injectablepharmaceutical compositions containing hydralazine and reduce thediscoloration of the hydralazine solution when diluted with conventionalpharmaceutical diluents.

It is a further object of the present invention to providepharmaceutical compositions containing N-protected hydralazine anddihydralazine derivatives in a formulation that is not metabolized byacetylation of the hydrazine group after administration.

DETAILED DESCRIPTION OF THE INVENTION

Compounds in accordance with the one embodiment of the present inventioninclude those containing a non-toxic, biocompatible N-protecting groupon the highly reactive hydrazine group of hydralazine represented by theformula:

where R₁ and R₂ are independently H, substituted or unsubstitutedbranched or straight chain alkyl having from 1 to about 7 carbon atoms,substituted or unsubstituted aryl, substituted or unsubstitutedcycloalkyl, substituted or unsubstituted aralkyl, substituted orunsubstituted alkylcycloalkyl, lower alkenyl or R₁ and R₂ together formpart of a substituted or unsubstituted cycloalkyl having from about 4 toabout 7 carbon atoms.

In one aspect of the present invention, R₁ and R₂ are preferablyunsubstituted branched or straight chain lower alkyls including but notlimited to methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl,isopentyl and hexyl groups. In another aspect of the present invention,R₁ and R₂ are substituted with hydroxyls. In this embodiment, R₁ is Hand R₂ has the formula CH₂(CHOH)_(m)CH₂OH where m is 2 or 3.

In one embodiment of the present invention, R₁ and R₂ are both branchedor straight chain lower alkyls. In another embodiment of the presentinvention, R₁ is a substituted or unsubstituted aryl, substituted orunsubstituted cycloalkyl, substituted or unsubstituted aralkyl,substituted or unsubstituted alkylcycloalkyl and R₂ is H or lower alkyl.In yet another embodiment of the present invention, R₁ and R₂ are bothsubstituted or unsubstituted aryl, substituted or unsubstitutedcycloalkyl, substituted or unsubstituted aralkyl, substituted orunsubstituted alkylcycloalkyl, lower alkenyl or R_(1 and R) ₂ togetherform part of a substituted or unsubstituted cycloalkyl having from about4 of to about 7 carbon atoms.

The preparation of acetone, 1-phthalazinylhydrazone is described in U.S.Pat. No. 2,484,029 issued on Oct. 11, 1949, which is hereby incorporatedby reference; however, no information regarding its stability in aqueoussolution is provided.

Compounds in accordance with this embodiment of the present inventionare readily prepared by reaction of the carbonyl group of the desiredacetone or aldehyde with the highly reactive primary amino group ofhydralazine (1-phthalazinylhydrazine). The resulting derivative ofhydralazine is generally called a hydrazone. Although aldehydes andketones are widespread in nature and are generally non-toxic, inpreferred embodiments of the present invention these aldehydes andketones will eventually end up in the patient's plasma after beingreleased from the hydralazine parent compound. Preferably, therefor, thealdehydes and ketones are non-toxic and biocompatible, and do not causeany deleterious effects in animals. Certain aldehydes and ketones havealready been recognized as less toxic and of lower risk to human healthby the FDA and are referred to as Class 3 compounds. These so-calledClass 3 compounds include those not known as a human health hazard atlevels normally accepted in pharmaceuticals even though there are nolong-term toxicity or carcinogenicity studies for many of the compounds.Available data indicate that they are less toxic in acute or short-termstudies and negative in genotoxicity studies. It is considered thatsmall amounts of these compounds in the amount of 50 mg per day or less(corresponding to 5,000 ppm would be acceptable without justification.

These hydrazone forming reactions are generally catalyzed by a smallamount of acid, and are buffered to a pH of about 4 to 5. In agreementwith general acid catalysis in which the conjugate acid of the carbonylreactant combines with a free amino group, the rate at which thesecompounds are formed generally drops at higher and lower pH values. Athigh pH there will be a vanishingly low concentration of the carbonylconjugate acid, and at low pH most of the amine reactant will be tied upas its ammonium conjugate acid. In general, these types ofderivatization reactions do not require active removal of water, and theproducts often precipitate from solution as they are formed.

In one embodiment of the present invention, hydralazine derivatives areprepared by coupling an N-protecting group to the terminal nitrogen ofthe highly reactive hydrazino group on hydralazine to produce a compoundhaving the formula:

where R₁ and R₂ are independently H, lower alkyl, or lower alkenyl.Because the hydrazine group readily reacts with the carbonyl group ofacetone and aldehydes the N-protected compounds of the present inventioncan easily be prepared by reacting hydralazine hydrochloride withaldehydes such as formaldehyde and acetaldehyde (I) or ketones such asacetone and other lower alkyl ketones such as butanone (II) asillustrated generally below:

Illustrative compounds in accordance with this embodiment of the presentinvention are found in Table 1.

TABLE 1 Ketones and Aldehydes Compound Number R₁ R₂ Starting Material  1H H formaldehyde  2 CH₃ H acetaldehyde  3 CH₃ CH₃ acetone  4 CH₂ CH₃ Hpropionaldehyde  5 CH₂ CH₃ CH₃ 2-butanone  6 CH₂ CH₃ CH₂ CH₃ 3-pentanone 7 CH₂ CH₃ CH₂CH₂ CH₃ 3-hexanone  8 CH₂CH₂ CH₃ H 1-butyraldehyde  9CH₂CH₂ CH₃ CH₃ 2-pentanone 10 CH₂CH₂ CH₃ CH₂CH₂ CH₃ 4-heptanone 11 CH(CH₃)₂ H 2-methyl propionaldehyde 12 CH (CH₃)₂ CH₃ 4 methyl butanone 13CH (CH₃)₂ CH₂ CH₃ 4 methyl 3-butanone 14 CH (CH₃)₂ CH₂CH₂ CH₃ 6-methyl4-hexanone 15 CH (CH₃)₂ CH (CH₃)₂ 2,4 dimethyl 3-pentanone 16 CH₂CH(CH₃)₂ H 3-methylbuteraldehyde 17 CH₂CH (CH₃)₂ CH₃ 4 methyl pentanone 18CH₂CH (CH₃)₂ CH₂ CH₃ 5 methyl hexanone 19 CH₂CH (CH₃)₂ CH₂CH₂ CH₃ 6methyl heptanone 20 CH₂CH (CH₃)₂ CH₂CH (CH₃)₂ 1-methyl, 7-methyl4-heptanone 21 CH₂CH₂CH₂ CH₃ H valeraldehyde 22 CH₂CH₂CH₂ CH₃ CH₃2-hexanone 23 CH₂CH₂CH₂ CH₃ CH₂ CH₃ 3-heptanone 24 CH₂CH₂CH₂ CH₃ CH₂CH₂CH₃ 4-octanone 25 CH₂CH₂CH₂ CH₃ CH₂CH (CH₃) CH₃ 2-methyl 4-octanone 26CH₂CH₂CH₂ CH₃ CH₂CH₂CH₂ CH₃ 5-nananone 27 CH₂CH₂CH₂ CH₂CH₃ CH₃2-heptanone 28 CH₂CH₂CH₂CH₂ CH₂CH₃ CH₃ 2-octanone 29 CH(CH₂CH₃)CH₂ CH₃CH₃ 3-ethyl 2-pentanone 30 CH(CH₃)CH₂ CH₃ CH₃ 3-methyl 2-pentanone 31CH(CH₃)CH(CH₃)CH₂ CH₃ H 2,3-dimethylpentaldehyde 32

CH₃ acetophenone 33

H benzaldehyde 34

CH₂CH₂CH₂CH₂ CH₂CH₃ 1-phenyl 1-heptanone 35

CH₂CH₂CH₂CH₂ CH₂CH₃ 1-phenyl 1-hexanone 36

H cyclohexanone 37

H 2-phenylacetaldehyde 38

CH₃ 3-phenyl 2-propanone 39

H 3-methyl 3-phenylbutyraldehyde 40

H 3-methyl 3- cyclohexylbutyraldehyde 41

CH₃ 4-methyl 4-phenyl 2-pentanone 42

CH₃ 4-cyclohexyl 4-methyl 2-pentanone 43

H p-methoxybenzaldehyde 44 CH₃ CH═C(CH₃)₂ 1,3-dimethyl-2-butenylidene 45H

46 H

TABLE 2 Compound Number R₁ and R₂ together Starting Material 47

cyclohexanone 48

4-methylcyclohexanone 49

cyclopentanone 50

3-methylcyclopentanone 51

cyclobutanone 52

3-methylcyclobutanone 53

cycloheptanone 54

3-methylcycloheptanone

In another embodiment, hydralazine derivatives are prepared by couplingan N-protecting group to the terminal nitrogen of the highly reactivehydrazine group on hydralazine to produce a compound having the formula:

where R₃ is a branched or straight chain alkyl having from 1 to about 7carbon atoms, substituted or unsubstituted aryl, substituted orunsubstituted aralkyl, substituted or unsubstituted cycloalkyl,substituted or unsubstituted aralkyl, substituted or unsubstitutedalkylcycloalkyl or a group having the formula (CH₂)_(n)COOH where n isfrom 1 to about 7. In accordance with one aspect of this embodiment ofthe present invention, R₃ is a branched or straight chain alkyl havingfrom 1 to about 7 carbon atoms. In accordance with another aspect ofthis embodiment of the present invention, R₃ is a substituted orunsubstituted aryl, substituted or unsubstituted aralkyl, substituted orunsubstituted cycloalkyl, or substituted or unsubstitutedalkylcycloalkyl.

Because the hydrazine group readily reacts with the carbonyl group ofacids the N-protected compounds of the present invention can easily beprepared by reacting hydralazine hydrochloride with acids such aspyruvic acid (III) α-ketoglutarate (IV) as illustrated generally below:

In accordance with a preferred embodiment of the present inventionhydralazine derivatives are prepared by coupling an N-protecting groupto the terminal nitrogen of the highly reactive hydrazine group onhydralazine to produce a compound having the formula:

where R₃ is a group having the formula (CH₂)_(n)COOH where n is from 1to about 4.

Illustrative compounds in accordance with this embodiment of the presentinvention are found in Table 3.

TABLE 3 Ketoacids Compound Number R₃ Starting Material 55 CH₃ Pyruvicacid 56 CH₂ CH₃ α-ketobutyric acid 57 CH₂CH₂ CH₃ α-ketopentanoic acid 58CH (CH₃)₂ α-ketoisovaleric acid 59 CH₂CH (CH₃)₂ α-ketoisocaproic acid 60CH₂CH(CH₃)CH₂ CH₃ 3-ethyl, 3 methyl pyruvic acid 61 CH₂CH₂CH₂ CH₃α-ketohexanoic acid 62 CH₂CH₂CH₂CH₂ CH₃ α-ketoheptanoic acid 63CH₂CH₂CH₂CH₂CH₂ CH₃ α-ketooctanoic acid 64

α-ketophenyl- pyruvic acid 65

α-ketophenyl- glyoxylic acid 66

α-keto 4-phenylbutyric acid 67

α-keto p-methoxyphenyl glyoxylic acid 68

4-methyl, 4-phenyl α-ketopentoic acid 69

α-ketocyclo- hexylglyoxylic acid 70 CH₂CH₂CH₂CH₂CH₂ CH₂CH₂COOHα-ketodecanedoic acid 71 CH₂CH₂CH₂CH₂ CH₂CH₂COOH α-ketononanedoic acid72 CH₂CH₂CH₂ CH₂CH₂COOH α-ketooctanoic acid 73 CH₂CH₂CH₂ CH₂COOH α-ketoheptanoic acid 74 CH₂CH₂CH₂COOH α-keto hexanoic acid 75 CH₂CH₂COOHα-ketoglutaric acid 77 CH₂COOH α-keto butanedioic acid 78 COOH α-ketoglyoxalic acid

Illustrative compounds in accordance with one embodiment of the presentinvention include the following:

Compounds in accordance with another embodiment of the present inventioninclude the following:

where X is a group having the formula

or compounds having the general formula:

where X is a group having the formula

where R₁, R₂ and R₃ having the same meaning as described above.

It will be apparent to those skilled in the art that compounds inaccordance with this embodiment of the present invention are similar tothose identified in Tables 1, 2 and 3 and can be prepared similarlyexcept that the starting material includes 1,4-hydrazinopthalazine(dihydralazine) instead of hydralazine.

In accordance with the present invention, stability with respect to thehydralazine compositions refers to both the chemical and physicalintegrity of the composition. Hydralazine hydrochloride, with a pK ofabout 7.3 is most stable at a pH of about 3.4 to about 4.4, and isunstable at high pH where it degrades into phthalazine, phthalazinoneand hydrazine. Because injectable hydralazine pharmaceutical solutionsare currently required to contain less than 10 ppm of hydrazine, sterileinjectable hydralazine solutions have a pH of between 3.4 and 4.4. Evenstored at a pH of about 3.4, sterile injectable hydralazine solutionsproduce detectable amounts of phthalazine, phthalazinone and hydrazineover time during storage. These sterile injectable hydralazine solutionsare filter believed to undergo degradation to insoluble polymericproducts through the highly reactive hydrazino group as illustratedbelow:

Degradation of Hydralazine on Storage

Although confirmation is not yet available, it is believed thatformation of the submicron particles in injectable hydralazine solutionsresults from the insoluble polymers of hydralazine generated throughdegradation during storage. In accordance with the present invention,the stability of hydralazine compositions during storage is determinedby monitoring the degradation products phthalazine, phthalazinone andhydrazine as well as the generation of insoluble polymeric products.Even stored at a pH of 3.4, sterile injectable hydralazine solutionsproduce detectable amounts of submicron particles over time duringstorage. In accordance with a most preferred embodiment of the presentinvention, the stability of the pharmaceutical compositions containinghydralazine is improved such that the shelf life of pharmaceuticalcompositions containing hydralazine is significantly extended beyond 12months storage and preferably beyond 24 months storage.

It is an object of the present invention that the stability of theN-protected hydralazine compounds is improved compared to that ofunconjugated hydralazine. The stability of the N-protected hydralazinecompounds of the present invention varies depending upon the nature ofthe protecting group; however, many of the compounds in accordance withthe present invention are acid-labile. Consequently, these compounds arestored at higher pH. In contrast to the sterile injectable hydralazinesolutions which are stored at a pH of about 3.4 to 4.4, the hydralazinecontaining pharmaceutical compositions in accordance with the presentinvention are preferable stored at a pH greater than 4.4. In a morepreferred embodiment of the present invention, the hydralazinecontaining pharmaceutical compositions of the present invention arestored at a pH of about 5 to about 8. The pH stability of the compoundsof the present invention is easily measured by testing solutions up to24 months using a HPLC method having a lower limit of sensitivity of0.0125 uM.

In a preferred embodiment of the present invention, stable hydralazinesolutions contain less than about 10 ppm (parts per million) and morepreferable less than about 3 ppm hydrazine. In a more preferredembodiment of the present invention, stable hydralazine compositionscontain less than about 1% by weight of degradation products includingphthalazine and phthalazinone. In a more preferred embodiment of thepresent invention, stable hydrazine solutions are essentiallyparticle-free. That is, the presence of particulate matter or particlesin injectable hydralazine compositions is not detectable by inspectingthe hydralazine solutions in both an upright and inverted position. In amost preferred embodiment of the present invention, the formation ofsubmicron particles in liquid pharmaceutical compositions containinghydralazine during storage is significantly reduced and particles arenot detectable from about 18 to about 24 months after completion ofmanufacturing and storage was initiated. In one embodiment of thepresent invention, injectable hydralazine formulations do not form smallyellow-green particles from 1 to about 2 months after storage whenhydralazine is stored at 40° C. and after from 6 to about 9 monthsstorage at 25° C.

In accordance with the present invention, particulate matter consists ofmobile randomly sourced extraneous substances, other than gas bubbles,that cannot be quantitated by chemical analysis due to the small amountof material that it represents and to its heterogeneous composition.Particulate matter (particles) in the injectable solutions forparenteral use in accordance with the present invention is determinedbased on visual inspection and by measured light obscuration procedures.Particles having a diameter of about 50 microns can be measured byvisual inspection. The light obscuration procedures are performed forthe purpose of enumerating subvisible extraneous particles having sizesthan about 50 microns. In accordance with the present invention,detection of particulate matter by light obscuration is preferablyperformed with a suitable electronic, liquid-borne particle countingsystem that uses a light-obscuration sensor with a suitablesample-feeding device. A variety of suitable devices of this type arecommercially available.

In accordance with the one embodiment of the present invention, aninjectable pharmaceutical composition is considered stable if theaverage number of particles of about 10 microns in the composition doesnot exceed 6,000. In accordance with another embodiment of the presentinvention, an injectable pharmaceutical composition is considered stableif the average number of particles of about 25 microns in thecomposition does not exceed 600. In accordance with yet anotherembodiment of the present invention, a pharmaceutical composition isconsidered stable if no particles are visible. In accordance with apreferred embodiment of the present invention, an injectable hydralazinecontaining pharmaceutical composition is particle-free if the averagenumber of particles of about 10 microns in the composition does notexceed 6,000, the average number of particles of about 25 microns in thecomposition does not exceed 600, and no particles are visible.

Acid labile derivatives of hydralazine have been reported by a number ofresearchers in an effort to identify and characterize the metabolites ofhydralazine including Clementi et al. in Journal of Pharmacological andExperimental Therapeutics 222(1): 159-165 (1982) found that certainacid-labile hydralazine derivatives were also plasma labile and arepharmacologically active and are endogenously hydrolyzed to parenthydralazine after intravenous administration. Clementi et al. reportthat, although differences in the pharmacological properties between thelabile derivatives related to the time course of parent hydralazinegeneration in plasma exist, the hydrolysis of the labile derivatives maybe nearly complete. Although differences in the extent and rate ofappearance of hydralazine in plasma are reported by Clementi et al., theextent and rate of appearance is therapeutically similar to that ofhydralazine after administration of hydralazine. Differences between thestability of hydralazine derivatives in plasma in vitro and the samecompounds in vivo suggest that plasma-labile derivatives of hydralazinemight be altered in the tissues as well as in the plasma.

The N-protected compounds produced from reaction with ketones, aldehydesor ketoacids in accordance with the present invention are used in thepreparation of a pharmaceutical dosage form intended for human use. Inthe case of manufacturing a sterile injectable dosage form suitable forintravenous administration to a patient, the N-protected compounds aredissolved in an appropriate solution for parenteral administration andfilled into bottles, vials, syringes or ampules with a pharmaceuticallyacceptable diluent under sterile manufacturing conditions. Uponcompletion of manufacturing this sterile injectable solution, the filledbottles, vials, syringes or ampules are stored under appropriate storageconditions. In the case of an oral dosage form, the N-protectedcompounds are mixed with pharmaceutically acceptable fillers andexcipients in a syrup, capsule or tablet.

In one embodiment of the present invention, the N-protected compoundsare formulated in a concentrated sterile solution for dilution at aconcentration of from about 10 to about 30 mg/ml (by weight), andpreferably at a concentration of about 20 mg/ml. Most preferably, thesecompounds are formulated in sterile water for injection at aconcentration of 20 mg/ml. In accordance with this embodiment of thepresent invention, the pH of the injection solution during storage isfrom about 7.4 to about 9.0, and preferably from about 8.0 to 8.5.

In another one embodiment of the present invention, the N-protectedcompounds are formulated in a more dilute concentration in a sterilesolution. In accordance with a preferred embodiment of the presentinvention, these compounds are formulated at a concentration of fromabout 0.5 to about 10 mg/ml (by weight), and preferably at aconcentration of about 5 mg/ml. Most preferably, these compounds areformulated in sterile water for injection at a concentration of 5 mg/ml.

After storage of the sterile injectable dosage form containing theN-protected compounds of the present invention, the N-protecting groupis removed from the parent hydralazine molecule immediately prior toinjection into a patient. In one embodiment of the present invention,the pH of the sterile injectable solution containing the acid-labilecompound is adjusted so that N-protecting group is released fromhydralazine and the N-protecting group remains in the injectionsolution. This is illustrated by the following reaction scheme:

By way of example, a vial containing 2 ml of the N-protected compoundsof the present invention formulated in a concentrated sterile solutionfor dilution at a concentration of about 20 mg/ml at pH 8.0, is dilutedprior to use with a sufficient volume of sterile water for injectionhaving a stabilizing effective pH. Preferably, the pH of the dilutedsolution is from about 3.0 to about 6.0.

Alternatively, the acid-labile compounds of the present invention areformulated into an oral dosage form such as a syrup, capsule or tablet.In this case, the syrup, capsules or tablets containing the N-protectingcompounds can be dosed to a patient without prior manipulation of thepH. In this case, as the oral dosage form reaches the highly acidicconditions of the stomach and the N-protecting group is removed from theparent hydralazine molecule prior to absorption. As in anotherembodiment of the present invention, the N-protecting group is releasedfrom hydralazine; however, the N-protecting group will be absorbed andmetabolized as it travels through the gastrointestinal system.

In the case of plasma-labile compounds of the present invention, thereis no need to adjust the pH of the composition so that N-protectinggroup is removed from the parent hydralazine compound. In this case, thecompounds of the present invention are administered to the patient andthe N-protecting group is removed in the plasma after administrationsuch that the extent and rate of appearance of hydralazine in the plasmais therapeutically similar to that of hydralazine after administration.In accordance with the present invention, the plasma-labile compoundsare therapeutically similar to hydralazine with regard to vasopressoractivity. It is contemplated that some differences in the extent andrate of appearance of hydralazine in the plasma between theseplasma-labile compounds and hydralazine will occur. Nevertheless, theirextent and rates of appearance in patients are to be consideredtherapeutically similar if these differences are medically insignificantor these compounds have the same clinical effect when administered topatients when administered under similar clinical conditions.

In yet another aspect of the present invention, the compounds areformulated in an oral pharmaceutical formulation and protected fromdegradation in the acidic environment of the stomach. This can beaccomplished, by way of example, through enteric coating of tablets orcapsules containing these compounds or by formulation of these compoundsin suitable drug delivery systems that release the pharmaceuticalcompounds only after the pharmaceutical preparation has passed throughthe stomach and entered into the small intestine. In this way, thecompounds of the present invention are presented to and adsorbed by theintestinal mucosa intact and are only metabolized to the hydralazinecompound during or after adsorption. This is particularly advantageouswhen administering hydralazine in an oral pharmaceutical composition topatients known as slow-acetylators, those having a deficiency ofN-acetyl transferase type 2 enzyme (NAT2). In accordance with thepresent invention, it is contemplated that the protective coatings willperform variably and might release a small portion of the compounds inthe stomach; however, a substantial portion of the pharmaceuticalformulation will only be released in the intestine. In a preferredembodiment of the present invention, no measurable amounts of thecompound are released from the dosage form in the acidic environment ofthe stomach and substantially all of the pharmaceutical compoundbypasses the stomach and is released in the intestine.

In accordance with the present invention and generally known to personsskilled in the art, a pharmaceutical compound is released from itsdosage form after dissolution or disintegration of structural componentsof the dosage form in a manner generally dependent upon its biologicalenvironment. For example, a compressed tablet or a capsule is capable ofdissolution and/or disintegration in the stomach; in which case, thedrug is released into the stomach. On the other hand, an enteric coatedtablet or capsule will resist degradation in the stomach and isessentially protected from degradation, (that is) it does not dissolveor disintegrate until it has transited to the more basic environment ofthe small intestine. Upon reaching the small intestine, the entericcoated tablet or capsule dissolves or disintegrates and its contents arereleased into the intestine.

In addition to the formation of hydralazine pyruvic acid hydrazone (HPH)the major plasma metabolite of hydralazine, hydralazine undergoes firstpass metabolism after oral administration by acetylation of thehydrazine group. This metabolic pathway, which is geneticallydetermined, occurs in the gastrointestinal mucosa and the liver by thetransfer of the acetyl group from acetyl coenzyme A to the hydrazinylgroup of hydralazine and the formation of the acetylation product(3-methyl-1,2,4-triazolo-(3,4a) phthalazine) known as MTP. MTP undergoesfurther metabolism by hydroxylation to form 3-hydroxymethyl-1,2,4-triazolo (3,4a) phthalazine (3-OHMTP). Hydralazine is alsometabolized by direct hydroxylation to 4-2-acetylhydrazinophhalazin-1-one (N-AcHPZ), which is mostly found in the urine.

N-acetyltransferase (NAT) is one of the major liver enzymes that areinvolved in biotransformation of drugs and other exogenous substances.It is recognized that biotransformation of xenobiotics containing anaromatic amine or a hydrazine group by N-acetylation is dependent on theN-acetyltransferases enzymes that in humans are expressed by only twodifferent enzymes, known as NAT1 and NAT2. Genetic polymorphismdetermines a reduction in the activity of the NAT2 enzyme that isobserved in slow acetylators. The incidence of slow acetylator phenotypeis about 5 to 10% in Asians, 50% in Americans, and 60 to 70% in NorthernEuropeans. NAT catalyzes the transfer of acetyl group from acetylcoenzyme A to the primary amino group of the acceptor molecule thatresults in the formation of N-acetyl derivatives. Individual variabilityin the activity of this enzyme was discovered nearly 40 years ago whenmarked interindividual variation in the elimination of isoniazid wasdetected. Further studies revealed that this variation was geneticallycontrolled and that slow acetylators of isoniazid were homozygous for arecessive gene, whereas rapid acetylators were either homozygous orheterozygous for the normal or wild type gene. Likewise, the metabolismof hydralazine after oral administration and formation of the acetylatedmetabolite is variable among individuals having phenotypic differencesin the expression of the NAT2 enzyme, the so-called fast-acetylators andslow-acetylators.

As a consequence of this difference in the metabolism of hydralazine,the fast-acetylators produce different amounts of active hydralazinemetabolites than the slow-acetylators, making it difficult to providethe correct dose of hydralazine to a patient in need of hypertensivetherapy. Moreover, a greater incidence of drug-induced lupuserythematosus has been associated slow-acetylators than withfast-acetylators after administration of hydralazine. The compounds ofthe present invention, when administered orally in a dosage form thatreaches the intestine intact, eliminate these differences. Becausesystemic metabolism is dependent on hydroxylation followed byconjugation with glucuronic acid in the liver and is not dependent onthe rate of acetylation, the half-life does not differ very much betweenslow and fast acetylators.

It is an object of the present invention to stabilize pharmaceuticalcompositions during manufacturing so that the hydralazine does not reactwith metal components of the manufacturing system. It is a furtherobject of the present invention to stabilize injectable pharmaceuticalcompositions containing hydralazine and reduce the discoloration of thehydralazine solution when diluted with conventional pharmaceuticaldiluents containing trace amounts of metals such as of Cu⁺², Fe⁺² andFe⁺³. Accordingly, the N-protected compounds of the present inventionhave a reduced capacity to complex with or otherwise react with metalsin the manufacturing solutions, storage solutions and diluent solutionsemployed in the hospital. In accordance with the present invention, thepharmaceutical compositions are essentially metal free and containessentially metal-free hydralazine. The presence of metals complexedwith hydralazine, as measured by the presence of color or reactivitywith spin-label probes, is a clear indication that the pharmaceuticalcompositions are not metal free and the compositions do not containmetal-free hydralazine.

EXAMPLE 1 Preparation of 1-hydrazinophthalazine

1-chlorophthalazine (30 parts) is heated for two hours in a mixture of100 parts by volume of ethyl alcohol and 90 parts by volume of hydrazinehydrate. After filtering, 1-hydrazinophthalazine crystallizes in yellowneedles on cooling. The yellow needles are filtered with suction andwashed with cold ethyl alcohol, and recrystalized from methyl alcohol(mp 172-178° C.). On warming in alcohol or aqueous hydrochloric acid,the hydrochloride is obtained (mp 273° C.).

EXAMPLE 2 Preparation of 1-hydrazinophthalazine α-ketoglutarateHydrazone

1-hydrazinophthalazine hydrochloride (395 mg; 2 mmol) is dissolved in 5ml of water. To this an aqueous solution of α-ketoglutaric acid (1 g; 7mmol) is added and the reaction mixture is allowed to stand overnight atroom temperature. The solid precipitate is filtered off and dried invacuo to yield 510 mg of I -hydrazinophthalazine a-ketoglutaratehydrazone (88% theoretical yield). [British Journal of ClinicalPharmacology 5: 489-494 (1978)]

EXAMPLE 3 Preparation of 1-hydrazinophthalazine Formaldehyde Hydrazone

Formaldehyde (0.3 g, 6.82 moles) is added with stirring to 500 ml of0.05 M phosphate buffer at pH 7.4 and 0.5 g (2.54 mmoles) of1-hydrazinophthalazine at 37° C. The reaction mixture is stirred at 37°C. for 10 min and then filtered. The filtrate is dried in vacuo to yielda solid residue. Recrystalization from chloroform-ether gives 0.440 g(92%) of the 1-hydrazinophthalazine formaldehyde hydrazone as off-whitecrystals, mp 108-110° C. [Journal of Pharmacological Sciences68(12):1524-1526 (1979)]

EXAMPLE 4 Preparation of 1-hydrazinophthalazine Acetaldehyde Hydrazone

Acetaldehyde (0.3 g, 6.82 moles) is added with stirring to 500 ml of0.05 M phosphate buffer at pH 7.4 and 0.5 g (2.54 mmoles) of I-hydrazinophthalazine at 37° C. The reaction mixture is stirred at 37°C. for 10 min and then filtered. The filtrate is dried in vacuo to yielda solid residue. Recrystalization from chloroform-ether gives 0.440 g(92%) of the 1-hydrazinophthalazine acetaldehyde hydrazone as off-whitecrystals, mp 108-110° C. [Journal of Pharmacological Sciences68(12):1524-1526 (1979)]

EXAMPLE 5 Preparation of 1-hydrazinophthalazine 2-butanone Hydrazone

A 10 parts 1-hydrazinophthalazine HCL (0.05 Molar) in 50% methanol-waterare mixed with 1 part 2-butanone. After evaporation of solvents,1-hydrazinophthalazine 2-butanone hydrazone is crystallized fromethanol-heptane (78% yield). [The Journal of Pharmacology andExperimental Therapeutics 205 (2): 418-425 (1978)].

EXAMPLE 6 Preparation of 1-hydrazinophthalazine Acetone Hydrazone

1-hydrazinophthalazine HCL (395 mg; 2 mmoles) is dissolved in 2.5 ml ofacetone and allowed to react for 1 hour. The solvent is evaporated andthe slightly yellow material is dried in vacuo to yield 411 mg of1-hydrazinophthalazine acetone hydrazone. (99% of theoretical yield).[British Journal of Pharmacology 61: 345-349 (1977)]

EXAMPLE 7 Preparation of 1-hydrazinophthalazine Pyruvate Hydrazone (a)

1-hydrazinophthalazine HCL (5 g; 25 mmoles) is dissolved in 50 ml 0.1 Msodium phosphate buffer pH 7.4, and a solution of sodium pyruvate (11 g;100 mmoles) in 30 ml of 0.1 M sodium phosphate buffer pH 7.4 is addedwhile stirring vigorously. The solution becomes distinctively yellowalmost immediately and the hydrazone is precipitated slowly. Afterstanding overnight at 4° C., the yellow crystalline product is filteredoff and washed with cold distilled water. The residue is recrystallizedfrom hot ethanol-water to yield 4.3 g of 1-hydrazinophthalazine pyruvatehydrazone (70% of theoretical yield). [Journal of Chromatography187:171-179 (1980)]

EXAMPLE 8 Preparation of 1-hydrazinophthalazine Pyruvate Hydrazone (b)

1-hydrazinophthalazine HCL (1 g) is mixed with acetone (0.38 g) in 30 mlwater at room temperature and the solution is magnetically stirred for10 minutes. Precipitated 1-hydrazinophthalazine acetone hydrazone iscollected by filtration as yellow crystals and dried over CaCl₂ [Journalof Pharmaceutical Sciences 85(3): 326-329 (1996)]

EXAMPLE 9 Preparation of 1-hydrazinophthalazine Anisaldehyde Hydrazone

1-hydrazinophthalazine HCL (395 mg; 2 mmoles) is dissolved in 2.5 ml ofanisaldehyde and allowed to react for 1 hour. The solvent is evaporatedand the slightly yellow material is dried in vacuo to yield 411 mg of1-hydrazinophthalazine anisaldehyde hydrazone. [Journal ofChromatography 126: 527-534 (1976)]

EXAMPLE 10 Single Dose Protocol in SHR [spontaneously Hypertensive Rats]

Adult male spontaneously hypertensive rats (SHR) weighing between 250and 350 g are maintained on standard rat chow and water ad libitum, andare prepared for cardiovascular studies by implantation of chronicaortic and jugular polyethylene catheters. Four days are allowed forrecuperation from surgery. Blood pressure is measured and recorded by aGrass model 7D polygraph (Grass Instrument Co., Quincy, Mass.) throughindividual Statham P23 Gb pressure transducers. An estimate of meanarterial pressure (MAP) is obtained by maximal electronic dampening ofthe input signal. During blood pressure measurements the animals areplaced in Plexiglas cages with minimal restraint. The MAP should rangefrom 150 to 170 mm Hg.

Each rat receives at least three different doses of i. v. bolusinjections of hydralazine and the compounds of Examples 2 through 8dissolved in a solution of sterile water for injection at pH 3.4. Drugadministrations are separated by 4 days to permit complete dissipationof drug effect. The volume of each injection is 0.1 ml/100 g by weightfor doses of 2.5 through 12.5 umol/kg. At doses of 20 umol/kg orgreater, the volume of each injection is increased to 0.2 ml/kg topermit complete solubilization of the N-protected hydralazine compounds.Hydralazine is dissolved in 30% EtOH-0.9% NaCl. After drugadministration, vasopressor activity is determined by measuring the meanarterial pressure (MAP) at 2, 4, 6, 8, 10, 15 and 30 min and then every30 min for 180 min. Dose response results are assessed by comparing theaverage peak change in MAP after administration at each dose of compoundadministered.

EXAMPLE 11 Stability of Hydralazine Hydrazones in Storage Solutions

The stability of the N-protected hydralazine compounds of the presentinvention is tested in solutions having a pH of 3.0, 4.0, 5.0, 6.0 7.0,8.0, 9.0 and 10.0 using a HPLC method having a lower limit ofsensitivity of 0.0125 uM. The amounts of hydrazine, hydralazine and theN-protected hydralazine compounds in an injectable formulation aremeasured using HPLC immediately after storing the solutions (T₀), 1week, 2 weeks, 1 month, 2 months, 3 months, 6 months, 9 months, 18months and 24 months after storage at 25° and 60° C. In addition, thestored solutions are visually inspected in both the upright and invertedpositions for the presence of particulate matter immediately afterstoring the solutions (T₀), 1 week, 2 weeks, 1 month, 2 months, 3months, 6 months, 9 months, 18 months and 24 months after storage at 25°and 60° C. The amounts of hydrazine, hydralazine and the N-protectedhydralazine compounds in an injectable formulation are also measuredusing HPLC immediately after storing the solutions (T₀), 1 week, 2weeks, 1 month, 2 months, 4 months, and 6 months after storage at 400 C.In addition, the stored solutions are visually inspected in both theupright and inverted positions for the presence of particulate matterimmediately after storing the solutions (T₀), 1 week, 2 weeks, 1 month,2 months, 4 months, and 6 months after storage at 40° C. The presence ofparticulate matter or small yellow-green particles over time is ameasure of stability.

EXAMPLE 12 Lability of Hydralazine Hydrazones in AdministrationSolutions

The lability of the N-protected hydralazine compounds of the presentinvention is tested in solutions having a pH of 3.0, 4.0, 5.0, 6.0 7.0,8.0, 9.0 and 10.0 using a HPLC method having a lower limit ofsensitivity of 0.0125 uM. After adjusting the pH of the storage solutioncontaining the N-protected hydralazine compounds, the hydrazones andhydralazine are measured at 10, 20, 30, 60 and 120 minutes.

The present invention has been described in detail using specificexamples to illustrate the preferred embodiments of the invention;however, it will be obvious to those skilled in the art that variousmodifications thereto can be made without departing from the spirit andscope thereof.

1. A compound having the formula:

or a compound having the formula:

where R₂ is H and R₁ is substituted cycloalkyl, substituted orunsubstituted aralkyl, or substituted or unsubstituted alkylcycloalkyl;where R₃ is substituted or unsubstituted aryl, substituted orunsubstituted aralkyl, substituted or unsubstituted cycloalkyl,substituted or unsubstituted alkylcycloalkyl or a group having theformula (CH₂)_(n)COOH where n is from 5 to about
 7. 2. A compound havingthe formula:

where R₃ is substituted or unsubstituted aryl, substituted orunsubstituted aralkyl, substituted or unsubstituted cycloalkyl,substituted or unsubstituted alkylcycloalkyl or a group having theformula (CH₂)_(n)COOH where n is from 4 to about
 7. 3. The compound ofclaim 2 wherein R₃ is substituted or unsubstituted aryl, substituted orunsubstituted aralkyl, substituted or unsubstituted cycloalkyl, orsubstituted or unsubstituted alkylcycloalkyl.
 4. The compound of claim 2wherein R₃ is a group having the formula (CH₂)_(n)COOH where n is from 4to about
 7. 5. A compound having the formula:

or a compound having the formula:

where R₁ and R₂ are independently H, branched or straight chain alkylhaving from 4 to about 7 carbon atoms, unsubstituted aryl, substitutedor unsubstituted aralkyl, substituted or unsubstituted alkylcycloalkyl;where R₃ is a branched or straight chain alkyl having from 4 to about 7carbon atoms, substituted or unsubstituted aralkyl, substituted orunsubstituted cycloalkyl, substituted or unsubstituted alkylcycloalkylor a group having the formula (CH₂)_(n)COOH where n is from 3 to about7; with the proviso that when R₁ is H then R₂ is substituted orunsubstituted alkylcycloalkyl.
 6. The compound of claim 5 wherein R₁ andR₂ are independently a branched or straight chain alkyl having from 4 toabout 7 carbon atoms, substituted or unsubstituted aralkyl, orsubstituted or unsubstituted alkylcycloalkyl.
 7. The compound of claim 5wherein R₂ is H and R₁ is a branched or straight chain alkyl having from4 to about 7 carbon atoms, unsubstituted aryl, substituted orunsubstituted aralkyl, or substituted or unsubstituted alkylcycloalkyl.8. The compound of claim 7 wherein R₁ and R₂ are a branched or straightchain alkyl having from 4 to about 7 carbon atoms.
 9. The compound ofclaim 7 wherein R₁ is a branched or straight chain alkyl having fromabout 4 to 7 carbon atoms.
 10. The compound of claim 5 wherein saidcompound has the formula:

where R₃ is a branched or straight chain alkyl having from 4 to about 7carbon atoms, substituted or unsubstituted aralkyl, substituted orunsubstituted cycloalkyl, substituted or unsubstituted alkylcycloalkylor a group having the formula (CH₂)_(n)COOH where n is from 3 to about7.
 11. The compound of claim 10 wherein R₃ is a branched or straightchain alkyl having from 4 to about 7 carbon atoms, substituted orunsubstituted aralkyl, substituted or unsubstituted cycloalkyl, orsubstituted or unsubstituted alkylcycloalkyl.
 12. The compound of claim10 wherein R₃ is a group having the formula (CH₂)_(n)COOH where n isfrom 3 to about
 7. 13. The compound of claim 5 wherein R₁ is anunsubstituted aryl, substituted or unsubstituted cycloalkyl, substitutedor unsubstituted aralkyl, or substituted or unsubstitutedalkylcycloalkyl.
 14. A compound having the formula:

where R₁ and R₂ are a branched or straight chain alkyl having from about4 to 7 carbon atoms.